Hazards and their exploitation in the applications of molecular biology to structure-function relationships.
Hazards and their exploitation in the applications of molecular biology to structure-function relationships.
复制标题
分子生物学在结构-功能关系中的应用中的危害及其利用。
DOI：
10.1021/bi00493a001
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发表时间：
1990
期刊：
影响因子：
2.9
通讯作者：
Schimmel,P
中科院分区：
文献类型：
--
作者：
Schimmel,P
分子生物学在酶学、蛋白质和核酸结构-功能关系方面的应用并不总是直截了当的。蛋白质和核酸的结构-功能关系的早期研究利用化学修饰和亲和标记来将功能分配给特定的氨基酸或核苷酸碱基，并使用有限的蛋白质水解或核酸酶消化来产生编码结构域的片段。通常，产生改变的核酸或蛋白质的操作以及改变的功能后果的分析都是在体外进行的。重组DNA技术有助于任何所需的核苷酸或氨基酸取代，并且原则上有助于任何所需片段的产生。然而，改变的分子的产生和至少一些它们的功能分析需要体内系统。在这种情况下，大量不完全表征或未知的变量可能导致结构和活动中的意外和未预料到的变化。这些并发症包括宿主细胞编码的分子和质粒编码的分子之间的新关联，质粒和宿主基因组序列之间的重组，其产生质粒编码的分子的序列变体，对在蛋白质或核酸片段的产生中暴露的序列的活性的特异质效应，以及密码子替换的翻译误读。在研究由重组分子表达的产物时，未能分析这些和其他可能性可能导致严重的解释错误。与此同时，这些现象中的一些现在已经被充分理解，它们可以被利用来进一步推进对结构-功能-活性关系的理解。遗传背景对蛋白质分析的意义
The applications of molecular biology to enzymology and to protein and nucleic acid structure-function relationships are not always straightforward. Early work on structure-function relationships of proteins and nucleic acids utilized chemical modifications and affinity labeling toassign functions to specific amino acids or nucleotide bases and used limited proteolysis or nuclease digestions to generate fragmentsthat encoded structural domains. Typically, manipulations that yielded altered nucleic acids or proteins, and analyses of functional consequences of alterations, were done invitro. Recombinant DNA technology facilitates any desired nu-cleotide or amino acid substitution and, in principle, production of any desired fragment. However, the production of altered molecules and at least some of their functional analyses re-quires in vivo systems. Here a large set of incompletely characterized or unknown variables can lead to unintended and unsuspected changes in structures and activities. These complications include novel associations between host-cell-encoded and plasmid-encoded molecules, recombination between plasmid and host genome sequences that generate sequence variants of the plasmid-encoded molecules, idiosyncratic effects on activity of sequences that are exposed in the creation of protein or nucleic acid fragments, and translational misreading of codon replacements. Failure to analyze for these and other possibilities in the investigation of products that are expressed from recombinant molecules can lead to serious errors of interpretation. At the same time, some of these phenomena are now sufficiently wellunderstood that they can be exploited to advance further the understanding of structure-function-activity relationships. Significance of Genetic Backgrounds for Protein Analysis